Recent studies indicate that the
chelator lipid nitrilotriacetic acid ditetradecylamine (
NTA-DTDA) can be used to engraft T cell costimulatory molecules onto
tumor cell membranes, potentially circumventing the need for genetic manipulation of the cells for development of cell- or membrane-based
tumor vaccines. Here, we show that a related
lipid 3(nitrilotriacetic acid)-ditetradecylamine (NTA(3)-DTDA, which has three NTA moieties in its headgroup instead of one) is several-fold more effective than
NTA-DTDA at promoting stable His-tagged
protein engraftment. IAsys biosensor studies show that binding of His-tagged B7.1 (B7.1-6H) to NTA(3)-DTDA-containing membranes, exhibit a faster on-rate and a slower off-rate, compared to membranes containing
NTA-DTDA. Also, NTA(3)-DTDA-containing
liposomes and plasma membrane vesicles (PMV) engrafted with B7.1-6H and CD40-6H exhibit greater binding to T cells, in vitro and in vivo. Engrafted NTA(3)-DTDA-containing PMV encapsulated
cytokines such as
IL-2,
IL-12,
GM-CSF and IFN-gamma, allowing targeted delivery of both
antigen and
cytokine to T cells, and stimulation of
antigen-specific T cell proliferation and cytotoxicity. Importantly, use of B7.1-CD40-engrafted PMV containing
IL-2 and
IL-12 as a
vaccine in DBA/2J mice induced protection against challenge with syngeneic
tumor cells (P815 mammary
mastocytoma), and regression of established
tumors. The results show that stable
protein engraftment onto liposomal membranes using NTA(3)-DTDA can be used to simultaneously target associated
antigen, costimulatory molecules and
cytokines to T cells in vivo, inducing strong anti-
tumor responses and immunotherapeutic effect.